Identification of Pathogenic Fusarium spp. Responsible for Root Rot of Angelica sinensis and Characterization of their Biological Enemies in Dingxi, China

Root rot is a serious disease in plantations of A. sinensis, severely affecting yield and quality and threatening sustainable production. Fusarium isolates (n=32) were obtained from field samples of root rot tissue, leaves and infected soil. Isolates were identified by comparing the sequences of their internal transcribed spacer (ITS) region and translation elongation factor 1-ɑ (TEF-1ɑ) to sequences of known species in the NCBI-database. These Fusarium isolates include F. tricinctum (43.75%), F. equiseti (31.25%), F. solani (9.37%), F. oxysporum (6.25%), F. acuminatum (6.25%), and F. incarnatum (3.12%). For pathogenicity testing under greenhouse conditions, seven isolates were selected based on a phylogenetic analysis, including four strains of F. tricinctum and one strain each of F. solani, F. oxysporum, and F. acuminatum. The seven isolates were all pathogenic but differed in their ability to infect: the four F. tricinctum strains were capable pathogens causing root rot in A. sinensis at 100% incidence and the highly aggressive. Furthermore, the symptoms of root rot induced by those seven isolates were consistent with typical root rot cases in the field, but their disease severity varied. Observed histopathological preparations of F. tricinctum-infected seedlings and tissue-slides results showed this fungal species can penetrate epidermal cells and colonize the cortical cells where it induces necrosis and severe plasmolysis. Plate confrontation experiments showed that isolated rhizosphere bacteria inhibited the Fusarium pathogens that cause root rot in A. sinensis. Our results provide timely information for informing the use of biocontrol agents for suppression of root rot disease.

Colonization of mutualistic mycorrhizal and parasitic blast fungi requires OsRAM2-regulated fatty acid biosynthesis in rice

Arbuscular mycorrhizal (AM) fungi form a mutual association with the majority of land plants, including most angiosperms of the dicotyledon and monocotyledon lineages. The symbiosis is based upon bidirectional nutrient exchange between the host and symbiont that occurs between inner cortical cells of the root and branched AM hyphae called arbuscules that develop within these cells. Lipid transport and its regulation during the symbiosis have been intensively investigated in dicotyledon plants, especially legumes. Here, we characterize OsRAM2 and OsRAM2L, homologs of M. truncatula RAM2, and found that plants defective in OsRAM2 were unable to be colonized by AM fungi and showed impaired colonization by Magnaporthe oryzae. The induction of OsRAM2 and OsRAM2L is dependent on OsRAM1 and the CSSP pathway genes CCaMK and CYCLOPS, while overexpression of OsRAM1 results in increased expression of OsRAM2 and OsRAM2L. Collectively, our data show that the function and regulation of OsRAM2 is conserved in monocot and dicot plants and reveals that, similar to mutualistic fungi, pathogenic fungi have recruited RAM2-mediated fatty acid biosynthesis to facilitate invasion.

Neuronal Activity Patterns Regulate Brain-Derived Neurotrophic Factor Expression in Cortical Cells via Neuronal Circuits

During development, cortical circuits are remodeled by spontaneous and sensory-evoked activity via alteration of the expression of wiring molecules. An intriguing question is how physiological neuronal activity modifies the expression of these molecules in developing cortical networks. Here, we addressed this issue, focusing on brain-derived neurotrophic factor (BDNF), one of the factors underlying cortical wiring. Real-time imaging of BDNF promoter activity in organotypic slice cultures revealed that patterned stimuli differentially regulated the increase and the time course of the promoter activity in upper layer neurons. Calcium imaging further demonstrated that stimulus-dependent increases in the promoter activity were roughly proportional to the increase in intracellular Ca2+ concentration per unit time. Finally, optogenetic stimulation showed that the promoter activity was increased efficiently by patterned stimulation in defined cortical circuits. These results suggest that physiological stimulation patterns differentially tune activity-dependent gene expression in developing cortical neurons via cortical circuits, synaptic responses, and alteration of intracellular calcium signaling.

Compartment-specific total RNA profile of Hippocampal and Cortical cells from Mesial Temporal Lobe Epilepsy tissue

Mesial temporal lobe epilepsy (mTLE) is a chronic neurological disease characterized by recurrent seizures. The pathogenic mechanisms underlying mTLE involve defects in post-transcriptional regulation of gene expression. So far, transcriptome profiles from epileptic tissue have been generated using whole cells, thereby lacking information on RNA localization and function at a subcellular level. In line with this, we have previously observed by in situ hybridization that a few microRNAs (miRNAs) display subcellular mis-localization with aberrant enrichment in the nucleus in human hippocampal mTLE tissue samples (Kan et al., 2012). To further investigate the possible mechanisms leading to the mis-localization of miRNAs, we set out to understand the compartment-specific total RNA (coding and non-coding) profile of human mTLE tissue samples. For this, we have successfully established a protocol to isolate cytoplasmic and nuclear compartments from human hippocampal tissue. After confirming the purity of the isolated cell compartments, we performed total RNA-sequencing (RNA-seq) on five resected hippocampal (HC) mTLE (no hippocampal sclerosis (non-HS)) samples and five HC postmortem control samples. Similarly, six neo-cortical (Cx) tissue samples from mTLE non-HS and HS International League Against Epilepsy (ILAE) Type 1, or mTLE+HS, samples were compared with six Cx postmortem controls. Our dataset provides a comprehensive overview of compartment-specific transcriptomic profiles of pharmacoresistant mTLE patient HC and Cx tissue, which in further studies can be used to investigate disease mechanisms.

An ATP-Binding Cassette Transporter, LaABCB11, Contributes to Alkaloid Transport in Lycoris aurea

As a kind of Amaryllidaceae alkaloid which is accumulated in the species of Lycoris plants, lycorine has a range of physiological effects. The biosynthesis pathway of lycorine has been partly revealed, but the transport and accumulation mechanisms of lycorine have rarely been studied. In this study, an ATP-binding cassette (ABC) transporter from Lycoris aurea (L’Hér) Herb., namely LaABCB11, was cloned and functionally characterized. Heterologous expression showed that LaABCB11 transported lycorine in an outward direction, increased the tolerance of yeast cells to lycorine, and caused a lower lycorine accumulation in transformants than control or mutant in yeast. LaABCB11 is associated with the plasma membrane, and in situ hybridization indicated that LaABCB11 was mainly expressed in the phloem of leaves and bulbs, as well as in the cortical cells of roots. These findings suggest that LaABCB11 functions as a lycorine transport and it might be related to the translocation and accumulation of lycorine from the leaves and bulbs to the roots.

Zn tolerance in the evergreen shrub, Aucuba japonica, naturally growing at a mine site: Cell wall immobilization, aucubin production, and Zn adsorption on fungal mycelia

Aucuba japonica Thunb. is an evergreen understory shrub that grows naturally at a mine site. The mine soil contains high concentrations of heavy metals, and A. japonica appears to maintain detoxification mechanisms against heavy metals in the study site’s understory. This study aimed to investigate the heavy metal tolerance mechanisms in A. japonica, considering the possible roles of arbuscular mycorrhizal and root-endophytic fungi. We conducted fieldwork in summer (canopy-foliation season) and winter (canopy-defoliation season) to measure the heavy metal concentrations in leaves, branches, and roots and analyze possible detoxicants in the roots. The infection rates of arbuscular mycorrhizal and root-endophytic fungi were evaluated via microscopic observation, and heavy metal (Zn) localization in A. japonica roots was observed using confocal laser scanning microscopy. Field analysis showed that A. japonica accumulated excessive Zn and produced aucubin and citric acid in the roots in both summer and winter. Zn localization observations clarified that Zn was distributed in thickened epidermal and cortical cell walls, suggesting that the cell walls functioned as Zn deposition sites, reducing Zn toxicity. It was further clarified that Zn was contained within cortical cells, indicating that Zn might be detoxified by aucubin and citric acid. Arbuscular mycorrhizal and root-endophytic fungi within cortical cells adsorbed Zn on fungal cell walls, indicating that these fungi would reduce Zn content within root cells and might alleviate Zn toxicity. Our results indicated that A. japonica would maintain Zn tolerance in both summer and winter via Zn immobilization in the cell walls and production of aucubin and citric acid, and that arbuscular mycorrhizal and root-endophytic fungi might play important roles in the Zn tolerance of A. japonica.

Microdevice for directional axodendritic connectivity between micro 3D neuronal cultures

Neuronal cultures are widely used in neuroscience research. However, the randomness of circuits in conventional cultures prevents accurate in vitro modeling of cortical development and of the pathogenesis of neurological and psychiatric disorders. A basic feature of cortical circuits that is not captured in standard cultures of dissociated cortical cells is directional connectivity. In this work, a polydimethylsiloxane (PDMS)-based device that achieves directional connectivity between micro 3D cultures is demonstrated. The device consists of through-holes for micro three-dimensional (μ3D) clusters of cortical cells connected by microtrenches for axon and dendrite guidance. The design of the trenches relies in part on the concept of axonal edge guidance, as well as on the novel concept of specific dendrite targeting. This replicates dominant excitatory connectivity in the cortex, enables the guidance of the axon after it forms a synapse in passing (an “en passant” synapse), and ensures that directional selectivity is preserved over the lifetime of the culture. The directionality of connections was verified morphologically and functionally. Connections were dependent on glutamatergic synapses. The design of this device has the potential to serve as a building block for the reconstruction of more complex cortical circuits in vitro.

Comprehensive in situ mapping of human cortical transcriptomic cell types

The ability to spatially resolve the cellular architecture of human cortical cell types over informative areas is essential to understanding brain function. We combined in situ sequencing gene expression data and single-nucleus RNA-sequencing cell type definitions to spatially map cells in sections of the human cortex via probabilistic cell typing. We mapped and classified a total of 59,816 cells into all 75 previously defined subtypes to create a first spatial atlas of human cortical cells in their native position, their abundances and genetic signatures. We also examined the precise within- and across-layer distributions of all the cell types and provide a resource for the cell atlas community. The abundances and locations presented here could serve as a reference for further studies, that include human brain tissues and disease applications at the cell type level.

Mixed corticomedullary tumor accompanied by unilateral aldosterone-producing adrenocortical micronodules: A case report

Mixed corticomedullary tumors (MCMTs) are rare and comprise of medullary and cortical cells in a single adrenal tumor. The mechanisms underlying its development have not been fully elucidated. Here, we report a case of MCMT in a 42-year-old woman. Based on the preoperative clinical findings, the patient was diagnosed as having a pheochromocytoma with subclinical Cushing's syndrome. Postoperative pathological diagnosis revealed that the tumor demonstrated morphologically distinct medullary and cortical components, which produced catecholamines and cortisol, respectively. Hybrid tumor cells producing both catecholamines and cortisol were not detected. Adrenocorticotropic hormone (ACTH)-positive tumor cells were identified to be present in the pheochromocytoma. This ectopic production of ACTH can contribute to an autonomous cortisol production in a paracrine manner. In addition, micronodules producing aldosterone were detected in the adrenal tissue adjacent to the tumor. The simultaneous development of these two lesions may not be correlated with each other; however, this case confirms the importance of a detailed histopathological examination of the adrenal lesions harboring complicated hormonal abnormalities by providing pivotal and indispensable information on their pathogenesis and the possible interaction of the hormones produced in the adrenal gland.